Chemokines are a large and growing family of 6-14 kD proteins that mediate a wide range of biological functions (Taub, D. D. and Openheim, J. J., Ther. Immunol., 1:229-246 (1994)). The chemokines can be divided into families based on the position of four cysteine residues that form two disulfide bonds (Kelner, G. S., et al., Science, 266:12395-1399 (1994); Bazan, J. F., et al., Nature, 385:640-644 (1997); Pin, Y., et al, Nature, 385:611-617 (1997)). Chemokine receptors can also be divided into families based on the type of chemokine they bind, although, no clear structural differences have been identified that distinguish the receptor sub-families (Mackay, C. R., J. Exp. Med., 184:799-802 (1996)). In addition, there are a number of so-called xe2x80x9corphanxe2x80x9d chemokine receptors which have amino acid sequences that are similar to well-characterized chemokine receptors, but for which the biological functions and specific receptor agonists (e.g., natural ligands) remain unknown.
Chemokines play a vital role in leukocyte adhesion and extravasation, and help direct lymphocyte localization to specialized microenvironments within tissues. Chemokines also help define sites of targeted lymphocyte trafficking from the blood. For example, the chemokines SLC and ELC which are both ligands for Cxe2x80x94C chemokine receptor 7 (CCR7) play a critical role in lymphocyte and dendritic cell trafficking to secondary lymphoid tissues, including lymph nodes (Warnock, R. A. et al., J. Exp. Med. 191:77 (2000); Stein J. V. et al., J. Exp. Med. 191:61 (2000); Iwasaki, A and Kelsall, B. L., J. Exp. Med. 191:1381 (2000); and Cyster, J. G., Curr. Biol. 10:R30 (2000)). Further, the chemokine TARC, a ligand for Cxe2x80x94C chemokine receptor 4 (CCR4), has been implicated in regulating lymphocyte-endothelial cell interactions and lymphocyte recruitment into non-intestinal tissues, particularly the skin (Campbell, J. J. et al., Nature 400:776 (1999)). Of further interest in the context of the regional specialization of immune responses is the constitutive and selective expression of chemokine TECK (a ligand for Cxe2x80x94C chemokine receptor 9 (CCR9)) by epithelial cells of the small intestine (Wurbel, M. A. et al., Eur. J. Immunol. 30:262 (2000)), and of the chemokine CTACK (a ligand for Cxe2x80x94C chemokine receptor 10 (CCR10)) by skin keratinocytes (Morales, J. et al, Proc. Natl. Acad. Sci. U.S.A. 96:14470 (1999)).
Agents which can inhibit particular chemokine/chemokine receptor interactions can prevent leukocyte trafficking (e.g., to particular tissues) and can have beneficial effects in models of acute and chronic inflammation ((Bargatze, R. F. and Butcher, E. C., J. Exp. Med., 178:367-372 (1993); Sekido, N., et al., Nature, 365:654-657 (1993); and Karpus, W. J., et al, J. Immunol., 155:5003-5010 (1995)). Therefore, there is a need for methods for identifying agents which can inhibit the binding of particular chemokines to particular chemokine receptors. There is also a need for methods for treating inflammatory conditions using such agents.
The invention relates to a method of detecting or identifying an agent (i.e., molecule or compound) which modulates the binding of a mammalian MEC to a mammalian Cxe2x80x94C chemokine receptor 3 (CCR3) and/or to a mammalian Cxe2x80x94C chemokine receptor 10 (CCR10) and/or which modulates the function of CCR3 and/or CCR10 upon MEC binding (e.g., MEC-induced signaling). Binding of agent to CCR3 and/or CCR10 can be detected directly or indirectly. For example, binding can be detected indirectly by monitoring a cellular response induced upon binding of MEC to CCR3 and/or CCR10. An agent which modulates (inhibits or promotes) the binding of MEC to CCR3 and/or CCR10 can be identified in a competitive binding assay. In some embodiments, the agent can be a small organic molecule. In other embodiments, the agent can be an antibody or antigen-binding fragment thereof. A suitable mammalian CCR3 or MEC-binding variant thereof and/or a mammalian CCR10 or MEC-binding variant thereof can be used. In one embodiment, a cell (e.g., primary cell, cell line, recombinant cell) that expresses a mammalian CCR3 or MEC-binding variant thereof is used. In another embodiment, a cell (e.g., cell line, recombinant cell) that expresses a mammalian CCR10 or MEC-binding variant thereof is used. In additional embodiments, a membrane preparation of a cell that expresses a mammalian CCR3 or MEC-binding variant thereof and/or a mammalian CCR3 or MEC-binding variant thereof is used.
The invention further relates to an agent, identified by the methods described herein, which modulates (inhibits or promotes) binding of a mammalian MEC to a mammalian CCR3 or MEC-binding variant thereof and/or a mammalian CCR10 or MEC-binding variant thereof and/or which modulates the function of mammalian CCR3 or variant and/or mammalian CCR10 or variant upon MEC binding (e.g., MEC-induced signaling). In one embodiment, the invention relates to an immunoglobulin or antigen-binding fragment thereof which binds a naturally occurring mammalian CCR10 and inhibits binding of a naturally occurring mammalian MEC to said receptor. In another embodiment, the invention relates to an immunoglobulin or antigen-binding fragment thereof which binds a naturally occurring mammalian MEC and inhibits binding of said MEC to a naturally occurring mammalian CCR3 and/or a naturally occurring mammalian CCR10. In a further embodiment, the invention relates to a small organic molecule which modulates (inhibits or promotes) binding of a mammalian MEC to a mammalian CCR3 or MEC-binding variant thereof and/or a mammalian CCR10 or MEC-binding variant thereof.
The invention also relates to therapeutic methods in which agents that modulate (inhibits or promotes) the binding of a mammalian MEC to a mammalian Cxe2x80x94C chemokine receptor 3 (CCR3) and/or to a mammalian Cxe2x80x94C chemokine receptor 10 (CCR10) are administered to a subject in need of such therapy. In one aspect, the therapeutic method is a method of treating a subject having an inflammatory disease. In one embodiment, the subject has an oral inflammatory condition, such as Sjogren""s syndrome or Behcet""s syndrome. In another embodiment, the subject has a chronic obstructive lung disease, such as asthma. In additional embodiments the subject has inflammatory bowel disease (e.g., Crohn""s disease, ulcerative colitis, celiac disease), mastitis, IgA nephropathy or dermatitis herpetiformis. In other embodiments, an immunoglobulin or antigen-binding fragment thereof which inhibits the binding of MEC to CCR3 and/or CCR10 is administered to a subject in need thereof.
The invention further relates to an agent which modulates (inhibits or promotes) binding of a mammalian MEC to a mammalian CCR3 or MEC-binding variant thereof and/or modulates binding of a mammalian MEC to a mammalian CCR10 or MEC-binding variant thereof (e.g., antibody, antigen-binding fragment), as described herein, for use in therapy (including prophylaxis) or diagnosis, and to the use of such an agent for the manufacture of a medicament for the treatment of a particular disease or condition as described herein (e.g., an inflammatory disease).
The invention also relates to an agent that modulates (inhibits or promotes) the activity of an IgA antibody-secreting cell in a subject. In one embodiment, the invention is a method of modulating the activity of an IgA antibody-secreting cell comprising administering an effective amount of MEC to a subject. In another embodiment, the invention is a method of modulating the activity of an IgA antibody-secreting cell comprising administering an effective amount of an agent that promotes the binding of MEC to CCR3 and/or CCR10 or an agent that inhibits the binding of MEC to CCR3 and/or CCR10.